Dataset Information

Rs10732516 polymorphism at the IGF2/H19 locus associates with genotype-specific effects on placental DNA methylation and birth weight of newborns conceived by assisted reproductive technology.

ABSTRACT: Background:Assisted reproductive technology (ART) has been associated with low birth weight of fresh embryo transfer (FRESH) derived and increased birth weight of frozen embryo transfer (FET)-derived newborns. Owing to that, we focused on imprinted insulin-like growth factor 2 (IGF2)/H19 locus known to be important for normal growth. This locus is regulated by H19 imprinting control region (ICR) with seven binding sites for the methylation-sensitive zinc finger regulatory protein (CTCF). A polymorphism rs10732516 G/A in the sixth binding site for CTCF, associates with a genotype-specific trend to the DNA methylation. Due to this association, 62 couples with singleton pregnancies derived from FRESH (44 IVF/18 ICSI), 24 couples from FET (15 IVF/9 ICSI), and 157 couples with spontaneously conceived pregnancies as controls were recruited in Finland and Estonia for genotype-specific examination. DNA methylation levels at the H19 ICR, H19 DMR, and long interspersed nuclear elements in placental tissue were explored by MassARRAY EpiTYPER (n?=?122). Allele-specific changes in the methylation level of H19 ICR in placental tissue (n?=?26) and white blood cells (WBC, n?=?8) were examined by bisulfite sequencing. Newborns' (n?=?243) anthropometrics was analyzed by using international growth standards. Results:A consistent trend of genotype-specific decreased methylation level was observed in paternal allele of rs10732516 paternal A/maternal G genotype, but not in paternal G/maternal A genotype, at H19 ICR in ART placentas. This hypomethylation was not detected in WBCs. Also genotype-specific differences in FRESH-derived newborns' birth weight and head circumference were observed (P?=?0.04, P?=?0.004, respectively): FRESH-derived newborns with G/G genotype were heavier (P?=?0.04) and had larger head circumference (P?=?0.002) compared to newborns with A/A genotype. Also, the placental weight and birth weight of controls, FRESH- and FET-derived newborns differed significantly in rs10732516 A/A genotype (P?=?0.024, P?=?0.006, respectively): the placentas and newborns of FET-derived pregnancies were heavier compared to FRESH-derived pregnancies (P?=?0.02, P?=?0.004, respectively). Conclusions:The observed DNA methylation changes together with the phenotypic findings suggest that rs10732516 polymorphism associates with the effects of ART in a parent-of-origin manner. Therefore, this polymorphism should be considered when the effects of environmental factors on embryonic development are studied.

SUBMITTER: Marjonen H

PROVIDER: S-EPMC6006593 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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rs10732516 polymorphism at the <i>IGF2/H19</i> locus associates with genotype-specific effects on placental DNA methylation and birth weight of newborns conceived by assisted reproductive technology.

Marjonen Heidi H Auvinen Pauliina P Kahila Hanna H Tšuiko Olga O Kõks Sulev S Tiirats Airi A Viltrop Triin T Tuuri Timo T Söderström-Anttila Viveca V Suikkari Anne-Maria AM Salumets Andres A Tiitinen Aila A Kaminen-Ahola Nina N

Clinical epigenetics 20180618

<h4>Background</h4>Assisted reproductive technology (ART) has been associated with low birth weight of fresh embryo transfer (FRESH) derived and increased birth weight of frozen embryo transfer (FET)-derived newborns. Owing to that, we focused on imprinted insulin-like growth factor 2 (<i>IGF2</i>)/<i>H19</i> locus known to be important for normal growth. This locus is regulated by <i>H19</i> imprinting control region (ICR) with seven binding sites for the methylation-sensitive zinc finger regul ...[more]

PMID: 29946374

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Project description:Study Question:Does prenatal alcohol exposure (PAE) affect regulation of the insulin-like growth factor 2 (IGF2)/H19 locus in placenta and the growth-restricted phenotype of newborns? Summary Answer:PAE results in genotype-specific trends in both placental DNA methylation at the IGF2/H19 locus and head circumference (HC) of newborns. What is Known Already:PAE can disturb development of the nervous system and lead to restricted growth of the head, even microcephaly. To clarify the etiology of alcohol-induced growth restriction, we focused on the imprinted IGF2/H19 locus known to be important for normal placental and embryonic growth. The expression of IGF2 and a negative growth controller H19 are regulated by the H19 imprinting control region (H19 ICR) with seven-binding sites for the methylation-sensitive zinc-finger regulatory protein CTCF. A single nucleotide polymorphism rs10732516 G/A in the sixth-binding site has shown to associate with genotype-specific DNA methylation profiles at the H19 ICR. Study Design Size Duration:By grouping 39 alcohol-exposed and 100 control samples according to rs10732516 polymorphism we explored alcohol-induced, genotype-specific changes in DNA methylation at the H19 ICR and the promoter region of H19 (H19 differentially methylated region). Also, IGF2 and H19 mRNA expression level in placenta as well as the phenotypes of newborns were examined. Participants/Materials Setting Methods:We explored alcohol-induced, genotype-specific changes in placental DNA methylation by MassARRAY EpiTYPER and allele-specific changes by bisulphite sequencing. IGF2 and H19 expression in placenta were analyzed by quantitative PCR and the HC, birthweight and birth length of newborns were examined using national growth charts. Main Results and the Role of Chance:We observed a consistent trend in genotype-specific changes in DNA methylation at H19 ICR in alcohol-exposed placentas. DNA methylation level in the normally highly methylated paternal allele of rs10732516 paternal A/maternal G genotype was decreased in alcohol-exposed placentas. In addition to decreased IGF2 mRNA expression in alcohol-exposed placentas of this specific genotype (P = 0.03), we observed significantly increased expression of H19 in relation to IGF2 when comparing all alcohol-exposed placentas to unexposed controls (P = 0.006). Furthermore, phenotypic examination showed a significant genotype-specific association between the alcohol exposure and HC of newborns (P = 0.001). Limitations Reasons for Caution:Owing to the exceptional character of the alcohol-exposed human samples collected in this study, the sample size is restricted. An increased sample size and functional studies are needed to confirm these data and clarify the biological significance or causality of the observed associations. Wider Implications of the Findings:Our results suggest that the rs10732516 polymorphism associates with the alcohol-induced alterations in DNA methylation profiles and head growth in a parent-of-origin manner. We also introduce a novel genotype-specific approach for exploring environmental effects on the IGF2/H19 locus and ultimately on embryonic growth. Study Funding/Competing Interests:This work was supported by the Academy of Finland (258304), The Finnish Foundation for Alcohol Studies, Finnish Cultural Foundation, Juho Vainio Foundation, Yrjö Jahnsson Foundation and Arvo and Lea Ylppö Foundation. No competing interests are declared.

Project description:BackgroundWith the increased use of assisted reproductive technology (ART), assessing the potential health risks of children conceived on ART important to public health. Most research in this area has focused on the effects of ART on perinatal, metabolic, and oncological risks in children. Although an increased risk of immune-related diseases has been reported in children born after ART, there are no studies on the immunological status of these children. This study aimed to evaluate the impact of different embryo transfer methods and fertilization strategies on the immune status of the offspring.MethodsA total of 69 children born to women treated with ART and a matched control group of 17 naturally conceived (NC) children, all aged from 3 to 6 years, were recruited in the reproductive hospital affiliated to Shandong University. The frequency of immune cells in the peripheral blood was assayed using flow cytometry; plasma cytokine levels were determined by multiplex cytokine immunoassay with human cytokine magnetic beads.ResultsCompared to children born after natural conception, children born after ART had elevated interferon-γ (IFN-γ) levels, regardless of embryo transfer and fertilization strategies. Children in the fresh-embryo transfer group had significantly higher IL-4 levels and a lower ratio of IFN-γ to IL-4 than those in the NC group ((P = 0.004, 10.41 ± 5.76 pg/mL vs 18.40 ± 7.01 pg/mL, P = 0.023, 1.00 ± 0.48 vs 0.67 ± 0.32, respectively). Similar results were shown in either the in vitro fertilization (IVF) group or the intra-cytoplasmic sperm injection (ICSI) group (P < 0.05 and P = 0.08 for IVF; P < 0.05 and P < 0.05 for ICSI, respectively). These alterations in IL-4 concentrations and the ratio of IFN-γ to IL-4 were statistically significantly correlated with supra-physical E2 (estradiol) levels on the day of hCG administration (R = 0.502, P = 0.017; R = - 0.537, P = 0.010, respectively). Consistently, the frozen embryo transfer did not result in alterations of these immune indicators in the offspring. Overall, there were no significant differences between the ART group and NC group in the frequencies of T cells, B cells, natural killer (NK) cells, CD4+T cells, CD8+T cells, T helper (TH)1 cells, TH17 cells, and regulatory T (Treg) cells and cytokine levels of IL-10 and IL-17a (all P > 0.05).ConclusionsImmunological alterations existed in children born after the use of ART. The elevated E2 levels before embryo implantation contributed to the increased IL-4 levels in children conceived by fresh embryo transfer. The assessment of immunological alteration is of importance to children conceived by ART for early monitoring and intervention.

Project description:Differentially methylated regions (DMRs) are associated with many imprinted genes. In mice methylation at a DMR upstream of the H19 gene known as the Imprint Control region (IC1) is acquired in the male germline and influences the methylation status of DMRs 100 kb away in the adjacent Insulin-like growth factor 2 (Igf2) gene through long-range interactions. In humans, germline-derived or post-zygotically acquired imprinting defects at IC1 are associated with aberrant activation or repression of IGF2, resulting in the congenital growth disorders Beckwith-Wiedemann (BWS) and Silver-Russell (SRS) syndromes, respectively. In Wilms tumour and colorectal cancer, biallelic expression of IGF2 has been observed in association with loss of methylation at a DMR in IGF2. This DMR, known as DMR0, has been shown to be methylated on the silent maternal IGF2 allele presumably with a role in repression. The effect of IGF2 DMR0 methylation changes in the aetiology of BWS or SRS is unknown.We analysed the methylation status of the DMR0 in BWS, SRS and Wilms tumour patients by conventional bisulphite sequencing and pyrosequencing. We show here that, contrary to previous reports, the IGF2 DMR0 is actually methylated on the active paternal allele in peripheral blood and kidney. This is similar to the IC1 methylation status and is inconsistent with the proposed silencing function of the maternal IGF2 allele. Beckwith-Wiedemann and Silver-Russell patients with IC1 methylation defects have similar methylation defects at the IGF2 DMR0, consistent with IC1 regulating methylation at IGF2 in cis. In Wilms tumour, however, methylation profiles of IC1 and IGF2 DMR0 are indicative of methylation changes occurring on both parental alleles rather than in cis.These results support a model in which DMR0 and IC1 have opposite susceptibilities to global hyper and hypomethylation during tumorigenesis independent of the parent of origin imprint. In contrast, during embryogenesis DMR0 is methylated or demethylated according to the germline methylation imprint at the IC1, indicating different mechanisms of imprinting loss in neoplastic and non-neoplastic cells.

Dataset Information

Rs10732516 polymorphism at the IGF2/H19 locus associates with genotype-specific effects on placental DNA methylation and birth weight of newborns conceived by assisted reproductive technology.

Publications

rs10732516 polymorphism at the <i>IGF2/H19</i> locus associates with genotype-specific effects on placental DNA methylation and birth weight of newborns conceived by assisted reproductive technology.

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